1. PackGene AAV helped the research of using AAVR to identify different mechanisms of AAV serotypes
Adeno-associated virus (adeno-associated virus, AAV) belongs to the Parvoviridae dependent virus genus. It has the advantages of good safety, wide tropism, can infect dividing or non-dividing cells, stable physical and chemical properties, and easy storage. It is widely used in gene therapy and vaccine research.
AAV receptors mainly refer to receptor molecules located on the surface of cell membranes that can specifically bind to AAV. The AAV receptor is the material basis for the specific recognition of AAV and target cells, and it is also the primary prerequisite for AAV to enter the cell and complete the life cycle in the cell. The AAVR protein is confirmed to be a kind of AAV receptor. AAVR is a transmembrane protein with immunoglobulin-like structure encoded by the KIAAO319L gene. It consists of a signal guide domain, an N-terminal transmembrane domain and an intracellular domain, and 5 Immunoglobulin-like extracellular domains (PKD1, PKD2, PKD3, PKD4, and PKD5) and a C-terminal transmembrane domain and intracellular domain are composed.
A paper entitled “Divergent engagements between adeno-associated viruses with their cellular receptor AAVR” was published by Tsinghua University School of Medicine in the internationally renowned academic journal “Nature Communications”, detailing the different mechanisms in the recognition of various AAV serotypes by AAVR. At the early stage, the research group used particle filter algorithm to analyze the high-resolution cryo-EM structure of the AAV2-AAVR complex with a resolution of 2.8Å. It was found that among the five PKD domains in AAVR, PKD2 directly binds to the spinous region of the AAV2 capsid adjacent to the icosahedral triplet axis, and the key binding site was determined.
In this study, the AAV virus and the packaging service were provided by PackGene.
2. PackGene AAV-CRISPR System helped the research on the Pathogenesis of Social Disorders in Autism
Positive Autism Spectrum Disorder (ASD) is a type of developmental behavior disorder characterized by varying degrees of social communication disorder, narrow interest, repetitive stereotyped behaviors, and abnormal perception. It seriously affects the life quality of patients and their families.
SHANK3 protein is the main scaffold protein of nerve synapses, distributed in the postsynaptic dense zone (PSD) of almost all excitatory synapses, and participates in the interaction between nerve transfer receptors and cytoskeleton proteins to form cytoskeleton-related signal complexes. It also participates in the formation and maturation of dendritic spines. Studies have found that the deletion, amplification, and mutation of the SHANK3 gene can cause the dysfunction of the SHANK3 protein, leading to various clinical symptoms of ASD.
An article titled Anterior Cingulate Cortex Dysfunction Underlies Social Deficits in Shank3Mutant Mice was published in Nature Neuroscience by Wu Shengxi and Wang Wenting’s team from the Department of Neurobiology, School of Basic Medical Sciences, Fourth Military Medical University, and Feng Guoping Laboratory of MIT. This article revealed the key role of the excitatory synapses of pyramidal neurons in the anterior cingulate cortex (ACC) in the social disorder of a mouse model of autism. The author alleviated or simulated social barriers by regulating the excitatory synapses of ACC pyramidal neurons forward or backward.This study provided the direct evidence for the causal relationship between ACC dysfunction and social disorder in Shank3 mutant ASD mouse model, emphasizing that ACC is a potential therapeutic target for social behavior deficit intervention.
In the study, CRISPR/Cas9 technology was used to knock out SHANK3 locally in ACC, and AAV was used as an efficient delivery system for gene knockout in vivo. The CRISPR-AAV virus used was packaged and produced by PackGene, and then injected into the mouse body through the orbital AAV. AAV is by far the safest viral vector which can infect the most wide range of tissues. PackGene combined AAV with the current revolutionary gene-editing CRISPR / systems to deliver gene knockout in vivo efficiently, and was applied in several recently published researches. The combination of AAV with strong tissue targeting and the CRISPR system that can perform multiple functions has become a powerful and potentially versatile animal experiment tool.
The gRNA backbones of all import vectors of PackGene have been optimized, which can increase the cutting efficiency by up to 40%. All CRISPR nuclease codes are optimized by mammalian/human codons. The CRISPR entry vector series (AA/AB/AC/AD/AE/AF/AG/AH) is a collection of the most cutting-edge CRISPR system achievements. Commonly used vectors have been verified by multiple cooperative units, which can greatly increase the experimental success rate.
3. The advantages of AAV produced by PackGene
Reliable
AAV packing titer is guaranteed over 1E+13GC/mL, and is quantified via ATCC Ref AAV. No worries for artificially high titer.
SDS-PAGE purity >95%, HPLC purity assay can be provided if requested
Tachypleus Amebocyte Lysate for endotoxin detection, ensure that the endotoxin level < 10EU/mL（1E+13GC）
Provide qPCR titer, SDS-PAGE purity and endotoxin assay reports for free (regular AAV packing)
Optional ddPCR, TEM empty capsid detection, mycoplasma detection, endotoxin removal assay and other services.
Provide 10% control AAV with high titer free ogre charge if needed（≥1E+13GC/mL）
Considerable
Customized AAV/ lentivirus packing services with guaranteed titer and quantity
QC report (protein purity examination, qPCR titer assay)
Deliver the AAV with dry ice with no delivery fee
Speed
Unique technology and projects management system
Deal with massive orders simultaneously with 95% orders being delivered within 6-9 workdays
Give 5% of delayed order sum as credit which can be used for future orders
Value
No worries for artificially high titer, quantity deficiency and other problems with relatively low price. PackGene will response as soon as possible if any problem occurs.
The link of AAV-CRISPR series product: https://www.packgene.cn/new_official_sub?category_id=2&sub_category_id=10&article_id=ed003621-c2b5-46a2-89db-5ab6926a510e
Reference:
https://www.sohu.com/a/329699207_650136 2. http://www.cnki.com.cn/Article/CJFDTotal-XZEK201404021.htm 3. http://www.cjebp.net/ CN/abstract/abstract9156.shtml
http://www.doc88.com/p-6992875626026.html
http://www.ddbjy.com/jiaoyu/gaoyi/152329.html
https://zhuanlan.zhihu.com/p/90254645
Anterior Cingulate Cortex Dysfunction Underlies Social Deficits in Shank3 Mutant Mice. Nat Neurosci. 2019 Aug;22(8):1223-1234. doi: 10.1038/s41593-019-0445-9. Epub 2019 Jul 22.
Divergent Engagements Between Adeno-Associated Viruses With Their Cellular Receptor AAVR . Nat Commun . 2019 Aug 21;10(1):3760. doi: 10.1038/s41467-019-11668-x.